Cow milk allergy [CMA] affects 2.5% of infants in the first 2 years of life or about 100,000 new cases per year in the U.S. IgE-mediated mechanisms account for 60% of these milk allergic disorders, with the majority involving the skin, while the majority of non-IgE-mediated reactions involve the gastrointestinal tract. About 80% of infants "outgrow" their CMA [develop clinical tolerance] in the first 3 - 4 years of life, but 35% of children with IgE-mediated CMA develop other food allergies and 60% develop respiratory allergy. Over the past granting period, we have enrolled a large cohort of well-defined patients with CMAs, compared humoral and cellular responses in different patient groups, and identified unique allergenic epitope recognition in patients with persistent CMA. However, the underlying milk-induced immunopathology of these disorders and their subsequent resolution remain poorly understood. Utilizing primary intestinal epithelial cell lines from different patient groups with CMA and normal controls, no differences were found in antigen processing including trafficking, cathepsin expression and activity, antigenic peptides, or the capacity to stimulate CD4 + or CD8 + T cell proliferation. Murine models of IgE-mediated CMA and isolated gut loops were developed to dissect immunoregulatory mechanisms involved in CMA and the role of the normal absorptive epithelium (E loops) vs the M cells and associated Peyer's patches in (M loops) in the development of tolerance. The combined resources of this program project provide a unique opportunity to define the immunologic bases for four common forms of CMA. Building on the well-defined patient cohorts enrolled in the program, the first project will further investigate unique humoral and cellular mechanisms underlying these disorders and changes associated with the development of clinical tolerance. The second project will focus on the function of intestinal epithelial cell CD23 as a bi-directional transporter of IgE and its role in CMA. The third project will utilize a novel in vivo gut-loop model to dissect immunologic mechanisms associated with the induction of normal gut-associated tolerance and pathologic responses of CMA. The fourth project will further investigate pathogenic immunoregulatory responses in murine models of CMA and cow milk tolerance, and evaluate the use of"engineered" recombinant proteins [from information gleaned in Project #1 ] to reverse CMA in mice with milk-induced anaphylaxis.